Antonio Raudino

α-Helical Structures Drive Early Stages of Self-Assembly of Amyloidogenic Amyloid Polypeptide Aggregate Formation in Membranes

The human islet amyloid polypeptide (hIAPP) is the primary component in the toxic islet amyloid deposits in type-2 diabetes. hIAPP self-assembles to aggregates that permeabilize membranes and constitutes amyloid plaques. Uncovering the mechanisms of amyloid self-assembly is the key to understanding amyloid toxicity and treatment. Although structurally similar, hIAPPs rat counterpart, the rat islet amyloid polypeptide (rIAPP), is non-toxic. It has been a puzzle why these peptides behave so differently. We combined multiscale modelling and theory to explain the drastically different dynamics of hIAPP and rIAPP: The differences stem from electrostatic dipolar interactions. hIAPP forms pentameric aggregates with the hydrophobic residues facing the membrane core and stabilizing water-conducting pores. We give predictions for pore sizes, the number of hIAPP peptides, and aggregate morphology. We show the importance of curvature-induced stress at the early stages of hIAPP assembly and the α-helical structures over β-sheets. This agrees with recent fluorescence spectroscopy experiments.

Hydrogen bonding and rotation barriers: A comparison between MNDO and AM1 results

Abstract Intra- and/or inter-molecular hydrogen bonding strengths, as well as rotation barriers, have been investigated for several molecular systems by means of MNDO and AM1 methods in order to test the reliability of the two semiempirical approaches. Comparison of the results show that although AM1 values do not satisfactorily match the experimental findings, they are far better than those of MNDO. In particular, the inability of MNDO to predict rotational barriers in conjugated molecules and hydrogen bonding energies has not been noticed in AM1.

Simulation of polyethylene glycol and calcium-mediated membrane fusion.

We report on the mechanism of membrane fusion mediated by polyethylene glycol (PEG) and Ca(2+) by means of a coarse-grained molecular dynamics simulation approach. Our data provide a detailed view on the role of cations and polymer in modulating the interaction between negatively charged apposed membranes. The PEG chains cause a reduction of the inter-lamellar distance and cause an increase in concentration of divalent cations. When thermally driven fluctuations bring the membranes at close contact, a switch from cis to trans Ca(2+)-lipid complexes stabilizes a focal contact acting as a nucleation site for further expansion of the adhesion region. Flipping of lipid tails induces subsequent stalk formation. Together, our results provide a molecular explanation for the synergistic effect of Ca(2+) and PEG on membrane fusion.

Inhibition of rat liver mitochondrial monoamine oxidase by hydrazine-thiazole derivatives: structure-activity relationships.

The purpose of this research is to study the relationship between chemical structure and inhibitory activity of some hydrazine-thiazole derivatives on rat liver mitochondria monoamine oxidase (MAO). Forty-five compounds belonging to three series of hydrazine-thiazole derivatives, with either alkylic or arylic substituents in the thiazole ring, were tested. The highest inhibitory activity was observed with piperonyl derivatives 25 and 40, which contain a 4-methyl group in the thiazole nucleus. The structure-activity relationship of MAO inhibitors was established in relation to hydrophobic, electronic and steric hindrance parameters. A mechanism of enzyme inhibition was proposed based on the calculation of HOMO energies.

MACROMOLECULAR PRODRUGS INTERACTION WITH MIXED LIPID-MEMBRANE - A CALORIMETRIC STUDY OF NAPROXEN LINKED TO POLYASPARTAMIDE INTERACTING WITH PHOSPHATIDYLCHOLINE AND PHOSPHATIDYLCHOLINE-PHOSPHATIDIC ACID VESICLES

Abstract The thermal behaviour of pure dipalmitoylphosphatidylcholine (DPPC) liposomes or mixed liposomes of DPPC with charged dipalmitoylphosphatidic acid (DPPA) and interacting with polymeric prodrugs has been investigated by differential scanning calorimetry (DSC). The apolar drug was naproxen (NAP) covalently linked to a water-soluble polymer (α,β- poly (N- hydroxyethyl )- dl - aspartamide (PHEA)). Addition of increasing amounts of NAP to DPPC liposomes causes a decrease in the transition temperature if( T m) associated to the gel-to-liquid crystal phase transition with a small decrease in the enthalpy values ( ΔH ), whereas a corresponding amount of drug contained in the PHEA-adduct modifies the liposome phase transition by decreasing the ΔH and broadening the peak without T m variations. These effects have been interpreted as a different interaction of free or polymer-bound drug with the lipid bilayer. The drug effect on mixed liposomes was also investigated, and evidence of improved interaction of the drug-PHEA adduct with two-component bilayers, which better mimic biological membranes, was found. In order to understand the prodrug-lipid interactions, we modulated the surface charge density of the mixed liposomes with Ca 2+ , which binds strongly to the negatively charged lipid head groups. We observed lateral phase separation, induced by NAP-PHEA adduct and modulated by Ca 2+ , and the phenomenon was explained in terms of different drug solubility in DPPC-poor and DPPC-rich microdomains. The results are indicative of different interactions of naproxen, either free or bound to a polymeric carrier, with phospholipid membranes and the ability of Ca 2+ to influence the adsorption of the drug.

Resveratrol interferes with the aggregation of membrane-bound human-IAPP: A molecular dynamics study

Amyloid aggregation of islet amyloid polypeptide (IAPP) in pancreatic tissues is a typical feature of type 2 diabetes mellitus. Resveratrol, a natural product extensively studied for its wide range of biological effects, has been shown to inhibit IAPP aggregation. However, the mechanism by which resveratrol inhibits IAPP aggregation is still far from complete elucidation. Now, an increasing knowledge of the mechanism of amyloid toxicity shifts the target of research towards the development of compounds which can prevent amyloid-mediated membrane damage rather than merely inhibit fiber formation. In this study we used all atom molecular dynamics to investigate the interaction of resveratrol with full-length human IAPP in a negatively charged membrane environment. Our results show that the presence of resveratrol induces the formation of secondary structures (sheets and helices) by inserting in a hydrophobic pocket between the interaction surface of two IAPP molecules in aqueous solution. On the other hand, resveratrol significantly perturbs the interaction of IAPP with negatively charged membranes by anchoring specific hydrophobic regions (23FGA25 and 32VGS34) of the peptide and forming a stable 1:2 IAPP:resveratrol complex at the water/membrane interphase.

Spectroscopic and molecular dynamics simulation studies of the interaction of insulin with glucose

The interaction between monomeric insulin and monosaccharides has been investigated through circular dichroism, fluorescence spectroscopy and two dimensional nuclear magnetic resonance. CD spectra indicate that D-glucose interacts with monomeric insulin whereas D-galactose, D-mannose and 2-deoxy-D-glucose have a lower effect. Fluorescence emission was quenched at sugar concentrations of 5-10 mM. Titration with the different sugars produces a quenching of the tyrosine spectrum from which a binding free energy value for the insulin-sugar complexes has been evaluated. Transfer nuclear Overhauser enhancement NMR experiments indicate the existence of dipolar interactions at short interatomic distances between C-1 proton of D-glucose in the beta form and the monomeric insulin. Further, NMR total correlation spectra experiments revealed that the hormone is in the monomeric form and that upon addition of glucose no aggregation occurs. The interaction does not involve relevant changes in the secondary structure of insulin suggesting that the interaction occur at the side chain level. Molecular dynamics simulations and modeling studies, based on the dynamic fluctuations of potential binding moiety sidechains, argued from results of NMR spectroscopy, provide additional informations to locate the putative binding sites of D-glucose to insulin.

Calcium ion influence on thermotropic behaviour of dipalmitoylphosphatidylcholine-vitamin D3 systems

An analysis of Ca2+ influence on the thermotropic behaviour of different phosphatidylcholine-vitamin D3 mixtures was carried out by differential scanning calorimetry technique (DSC). A competitive effect between Ca2+ and vitamin D3 on the membrane fluidity was detected. The observed shifts of the gel-liquid crystal transition temperature were correlated with the mole fraction of vitamin D3 dissolved within the lipid bilayer as well as with the Ca2+ concentration in the surrounding medium. These shifts were rationalized on the ground of a simple microscopic model through the calculation of the internal pressure exerted by the adsorbed Ca2+ on the lipid matrix by the Clapeyron equation. The experimental results and the obtained equations accord with each other and support the idea of micro-domain formation richer in one lipid component. The extent of such lateral phase separation of the lipid components seems to be favoured by the adsorption of Ca2+ at the membrane-water interface.

New interferometric technique to evaluate the electric charge of gas bubbles in liquids.

We report a new interferometric technique to measure the electric charge at the gas-liquid interface of a bubble in a liquid. The bubble rests by buoyancy against an electrode, and an alternating electric field excites its capillary oscillations. The oscillation amplitude of the quadrupolar mode frequency is measured by the interferometer, and it is used to evaluate the electric charge. The mode frequency scales with the square root of the interfacial tension and with a -(3)/(2) power law as a function of the bubble radius. For bubbles in the millimeter diameter range in pure water, the measured negative charge scales with the square of the radius, hence, giving a constant surface charge density on the order of 1.8 × 10(-5) C m(-2), which is rather consistent with the electrophoretic values reported in the literature.

Collective phenomena in confined micellar systems of gangliosides

In the present paper, we investigate in deeper detail some features of the bistable behavior observed in micellar aggregates of gangliosides. In these systems, a collective transition takes place connected to different possible conformations of the glycosidic headgroup and resulting in different aggregation numbers for different thermal hystories. Light scattering experiments performed on mixed micelles indicate that it is the aggregated structure which dictates the conformation a ganglioside monomer assumes, among the allowed ones. Moreover, density measurements confirm the results previously obtained by DSC and show that the conformation of the hydrophobic tails at a given temperature is coupled to that of the headgroup. NMR measurements are consistent with previous results and, on a more local scale, indicate that the hydration of the ganglioside molecule is affected by the conformational transition.